1. Field of the Invention
The present invention relates to high-pressure fluorescence flow cells, assemblies thereof, fluorescence detectors using the high-pressure fluorescence flow cells, and supercritical fluid chromatographs.
2. Description of the Related Art
Supercritical Fluid Chromatograph
Supercritical fluid chromatographs (SFCs) and subcritical fluid chromatographs using carbon dioxide (CO2) as the mobile phase have attracted attention as separation-analysis apparatuses and separation-purification apparatuses that can replace liquid chromatographs (LCs), which utilize an organic solvent. Thanks to the low viscosity and high diffusion coefficient of the fluid utilized as the mobile phase, SFCs can perform quick analysis by increasing the flow velocity without decreasing the column efficiency.
SFCs can be divided broadly into two categories depending on the type of column used. The SFCs utilizing a packed column use techniques developed for liquid chromatographs (LCs), and the SFCs utilizing a capillary column use techniques developed for gas chromatographs (GCs). Since packed-column SFCs developed from LCs are easy to handle and their separation technology is extensible, they have been used to analyze a variety of samples and have been finding a broader range of applications. SFCs are used to separate optical isomers and cc separate polymer oligomers in polymer chain units, are environmentally friendly, and have low running costs. SFCs require a few aftertreatment steps for eliminating the solvent and are suitable for easily-oxidizable material and heat-labile material. In recent studies, SFCs have found wide applications, including separation of achiral matter.
Such separation-analysis apparatuses have also attracted attention from the perspective of environmental conservation. There seems to a tendency to shift from liquid chromatographs (LCs) to supercritical fluid chromatographs (SFCs).
Detectors for Supercritical Fluid Chromatographs
Detectors used for supercritical fluid chromatographs (SFCs) include ultraviolet-visible (UV-VIS) absorbance detectors, evaporative light scattering detectors (ELSDs), and mass spectrometers (MSs). Some of the detectors, such as ultraviolet-visible absorbance detectors, require an optical flow cell. The flow cell for SFCs must have high pressure resistance because the inner pressure reaches 10 MPa or higher.
Fluorescence Detectors
Fluorescence detectors (FLDs) for detecting fluorescence (including phosphorescence) of excited sample constituents also require an optical flow cell, FLDs are used with liquid chromatographs (LCs). The conventional fluorescence detectors have a rectangular flow cell made of silica glass, for example. The sample constituents separated in the column of the LC are introduced successively into the flow cell together with a mobile phase For example, the sample constituents are introduced into the rectangular flow cell from the bottom, travel through an internal flow path, and are drawn out from the top of the flow cell. While they are in the flow path, the sample constituents are irradiated with excitation light from one side of the flow cell. When fluorescence is produced in the sample constituents, the fluorescence is detected in a direction perpendicular to the irradiation direction of the excitation light.
In general separation and analysis of a sample by using a chromatograph, it is important to reduce the volume of the cell to prevent the sample constituents separated in the column from diffusing in the flow cell. For example, low-volume flow cells have been formed by fusing four light-transmissive members made of silica glass into a hollow prismatic shape, as disclosed in Patent Literature 1. By minimizing the cell volume, diffusion of the sample constituents in the cell can be suppressed, and consequently peaks in the detected chromatogram can be sharpened.
In order to prevent the minimized cell volume from lowering the sensitivity of the fluorescence detector, a flow cell configured so allow internal sample constituents to be irradiated efficiently with as much excitation light as possible is used. If the sample is irradiated with excitation light in a direction perpendicular to the flow path of the cell, as the cross section of the flow path in the cell decreases, the optical path length of the cell inevitably decreases. For example, the flow cell disclosed in Patent Literature 2 is configured to direct excitation light parallel to the direction of the flow path. A long optical path length is kept to produce a large amount of fluorescence.